Derivation of authentic porcine embryonic stem cells using defined culture conditions

Pig embryonic stem cells (ESCs) have been considered as an important candidate for preclinical researches on human therapy. However, the lack of understanding of pig pluripotent networks has hampered establishment of authentic pig ESCs. Here, we report that FGF2, ACTVIN, and WNT signaling is essential for maintaining pig pluripotency in vitro. Pig ESC lines derived by stimulating three signlaings formed colonies of flattened monolayer morphology. Newly derived ESCs were stably maintained over an extended period, and were capable of forming teratomas comprising three germ layers. Immunostaining showed that the stem cells expressed pluripotency markers including OCT4, SOX2, NANOG, SSEA1, and SSEA4. Transcriptome analysis showed that pig ESCs were developmentally similar to late epiblasts of preimplantation embryos and in terms of biological functions resembled human rather than mouse ESCs. However, the pig ESCs had distinct features such as coexpression of SSEA1 and SSEA4, two active X chromosomes, and a unique transcriptional pattern. In conclusion, we successfully derived authentic pig ESCs using novel cell culture conditions. Our findings will facilitate both the development of large animal models for human stem cell therapy and the generation of pluripotent stem cells from other domestic animals for agricultural use.